1
Q
what makes up most of the cells dry mass?
A
-proteins
2
Q
what are proteins?
A
-main building blocks of the cell
3
Q
what are the different kinds/functions of proteins?
A
-enzymes (catalysts, ase = breaking)
-structural (provide support, ex: keratin)
-transport (carry small things across, membrane embeded)
-motor (propel things through the cytosol or tissue ex: myosin in muscle)
-storage (AA or ions)
-signalling (within a cell or btwn cells, relay info)
-receptors (detect signals + transmit them)
-transcription regulators (turn genes on/off)
-MANY MORE
4
Q
what does a dehydrogenase protein do?
A
-enzyme that removes e-
5
Q
what is important to remember with proteins?
A
-structure = function
6
Q
what is the basic structure of a protein?
A
-amino acids joined by covalent peptide bonds (form polypeptide chains)
-have a polypeptide backbone which is a repeating of core atoms (-N-C-C-)
-N terminus direction (carries an amino group)
-C terminus direction (carries a carboxyl group)
-side chain (R group/variable group)
7
Q
what is the N-terminus end of a polypeptide chain?
A
-end that carries the amino group
-typically present in ionized form due to the cell environment (NH3+ not NH2)
8
Q
what is the C-terminus end of a polypeptide chain?
A
-end that carries the carboxyl group
-typically present in ionized form due to the cell environment (COO- not COOH)
9
Q
what is the purpose of the C/N terminus ends of a polypeptide chain?
A
-give directionality
10
Q
what is the purpose of the side chain (R group/variable group) on each amino acid?
A
-gives the amino acid its identity and unique properties
-may have a charge (reactivity)
-can be polar (hydrophillic) or non-polar (hydrophobic) (charge)
11
Q
are the side chains involved in peptide bonding?
A
-NO
12
Q
what is important within polypeptide chain?
A
-the distribution of polar and non-polar amino acids (for shape)
13
Q
what are the features of polypeptide chains and their ability to form conformations?
A
-chains are flexible due to the rotation around single covalent bonds (allows for various folding)
-shape is constrained by weak interactions within the molecule (backbone and side chains)
14
Q
what determines the stability of a polypeptide molecule?
A
-the combined strength of all weak interactions (H bonds, VDW) and electrostatic interactions aka ionic bonds
15
Q
what is the hydrophobic force? what does this cause?
A
-the forcing of the non-polar side chains to be clustered together towards the inside of a folded protein
-causes an increase of VDW
16
Q
where are the polar amino acids found on a folded protein?
A
-near the outside typically (hydrophilic and want to interact with aqueous solutions such as cytosol)
-can sometimes be buried on the inside if they are hydrogen bonded to other polar amino acids
17
Q
what are backbone to backbone interations?
A
-a hydrogen bond between atoms of two peptide bonds
18
Q
what are backbone to side chain interactions?
A
-a hydrogen bond between atoms of a peptide bond and an amino acid side chain
19
Q
what are side chain to side chain interactions?
A
-a hydrogen bond between atoms of 2 amino acid side chains
20
Q
is protein folding energetically favourable or energetically unfavourable?
A
-energetically favourable
-the final conformation of a protein will always minimize the free energy and thus release heat and increase disorder of the universe
21
Q
what is conformation?
A
-the final folded structure of a polypeptide chain
22
Q
what is denaturation?
A
-protein losing its conformation due to the disruption of non-covalent bonds
-occurs due to a change in environment (pH or addition of a solvent like SDS)
-high energy
23
Q
what is renaturation?
A
-spontaneous refolding of a protein when the proper conditions are provided
-works best for small proteins
24
Q
what do protease enzymes act on?
A
-the peptide bonds between amino acids
-they will not worry about the weak interactions
25
what are chaperone proteins?
-proteins that assist a polypeptide to fold into its most energetically favourable conformation
-require ATP binding and hydrolysis
-increase the efficiency and reliability of folding
-tuck pieces by binding to partly folded chains
26
is the folding of a protein with the help of chaperone proteins fully spontaneous?
-since the chaperone proteins assist, folding is NOT fully spontaneous
27
what can chaperone proteins form?
-isolation chambers
28
what are isolation chambers?
-a set of chaperone proteins that form a cage where the protein can properly fold
-prevents the polypeptide from aggregating with other polypeptides (causes them to not form since other interactions get in the way)
29
when does the conformation of a protein change slightly?
-when a protein interacts with other molecules in the cell
-shape changes are CRUCIAL to function
30
what kinds of shapes can proteins form? what are their general size?
-size is 30-10000 amino acids (avg = 50-2000)
-form filaments, sheets, rings, spheres, globulars (basically any shape)
31
what are the different models to show protein shape?
-backbone model
-wire model
-ribbon model
-space-filling model
-choose one depending on what you are trying to show
32
what does the backbone model show?
-just shows the polypeptide backbone
-most simple and straightforward for comparison
33
what does the ribbon model show?
-shows the secondary structure (alpha helices and beta sheets)
-shows some folding patterns
34
what does the wire model show?
-shows side chains and backbone
-can see what is in the active site, charges, what amino acids are present (important for determining activity)
35
what does the space filling model show?
-surface AA (cannot see exactly which ones are fully exposed, but can see most)
-can see how it may look to other molecules
36
what is the protein HPr?
-bacterial transport protein that facilitates the transport of sugars into the cell
37
what is primary structure?
-amino acid seqeunce
38
what is secondary structure?
-alpha helices
-beta sheets
39
what is tertiary structure?
-full 3D conformation of the polypeptide chain
-folding in on itself due to the weak interactions
-can end here or reach one more organization level
40
what is quaternary structure?
-multiple polypeptides interacting to form the complete protein
-multiple subunits present held by non-covalent bonds
41
what are alpha helices?
-many similar subunits next to each other in the same repeated relationship to the one before
-hydrogen bonds between every 4th amino acid (C double bonded O of one peptide bond to the N-H bond of another)
-right handed helix with a complete turn every 3.6 amino acids (rare to end up in a straight line) (turns clockwise)
42
where are alpha helices abundant?
-in proteins that are embedded in the cell membrane (transporters or receptors)
-structure allows them to drill through the membrane to be transmembrane proteins (some protein on the outside of the membrane)
43
what type of amino acids are typically within transmembrane proteins?
-non polar amino acids (in the section that crosses the membrane)
-these AA side chains protrude out and the backbone is H bonded to itself inside the helices (to shield from the hydrophobic lipid environment)
44
what is a coiled coil?
-two or three alpha helices wrapped around eachother
-very stable structure (used for things that need a lot of strength)
-non polar side chains are typically along one side so they can twist around with their hydrophobic regions inwards to minimize cytosol contact (increased vdw + hydrophobic forces)
45
what proteins form coiled coils?
-elongated proteins such as alpha keratin, intermediate filaments and myosin
46
what are beta sheets?
-rigid structures at the core of proteins (form the bulk)
-hydrogen bonds between neighbouring segments of polypeptides
-can have 2 formations
47
what are the 2 formations of beta sheets?
-parallel beta sheets
-antiparallel beta sheets
48
what are parallel beta sheets?
-polypeptide segments run in the same direction
-ex: N-C, N-C, N-C
49
what are antiparallel beta sheets?
-polypeptide segments run in opposite directions
-ex: N-C, C-N, N-C, C-N
50
what can beta sheets form?
-amyloid structures
51
what are amyloid structures?
-beta sheets stacked together with side chains interdigitated like the teeth of a zipper
-used for the storage of peptide or protein hormones for efficient packaging in transport vesicles
-used in cells specialized for secretion (unfold once the exterior is reached)
-can be damaging in some cases
52
how can amyloid structures be damaging to the cell?
-misfolded proteins often form amyloid structures that damage cells (specifically in the brain)
-present in diseases such as alzheimers, parkinsons, and huntingtons disease which are all neurodegenerative (neurons cannot always regenerate)
-some amyloid proteins can move from cell to cell causing further misfolding (prion-like)
53
what are prions?
-infectious misfolded proteins which contain amyloid structures
-can convert normal proteins into adopting an amyloid structure (prion form)
-can be caused by mutation, rarely ever spontaneous (do not see many people with it)
-resistant to denaturing (must make a solution and then put in the autoclave)
54
what diseases are caused by prions?
-scrapie (sheep)
-bovine spongiform (mad cow disease)
-encephalopathy
-creutzfeldt-jakobs disease (human)
-chronic wasting disease (deer)
55
how long does it take for symptoms of creutzfeldt-jakobs disease to show?
-5-10 years
56
why does chronic wasting disease happen?
-if a deer does not have enough vegetation to eat, then it eats dead deer and spreads prions
-deer becomes unbalanced and lacks control (should not eat animals with these symptoms)
57
what is a protein domain?
-any segment of a polypeptide that can fold independently into a compact and stable structure (ex: alpha helices, beta sheets, other structure elements)
-typically 40-350 amino acids
-each domain has its own function (often)
-linked together by unstructured sequences (short polypeptide chains)
58
what is an example of a protein with multiple domains?
-bacterial transcriptional regulator (determines if transcription is going to occur)
-small domain that binds DNA
-large domain that binds cAMP (circularized AMP)
-when the large domain binds cAMP, it causes a conformational change in the small domain so that it can bind DNA
59
what are protein families?
groups of proteins that closely resemble each other
-each protein will still have its own distinct enzymatic function
-look similar but has enough difference to have a small function change
60
what is an example of a protein family?
-serine proteases
-elastase vs chymotrypsin
-elastase cleaves carboxyl groups on small hydrophobic AA's
-chymotrypsin hydrolyses peptide bonds between specific AA's
61
what is a binding site?
-region on a protein's surface that interacts non-covalently with another molecule
-proteins can each bind to many molecules
62
if a protein is binding to another protein what do we call each polypeptide chain?
-a subunit
-each subunit can have multiple domains
-ex: dimer, tetramer
63
what is a dimer?
two identical polypeptide chains (subunits) bound together (each has a binding site that is identical to the other)
64
what is a tetramer?
-four identical polypeptide chains (subunits) bound together
65
are subunits always identical to one another?
-NO
-subunits can also be non identical (still facilitate function)
-ex: hemoglobin (2 alpha globin SU, 2 beta globin SU) or RNA polymerase holoenzyme
66
what are examples of proteins that are chains of identical proteins often in the shape of a helix?
-actin filaments
-microtubules (each dimers of tublin, has sites to bind dimers together)
-all extended protein filaments
67
what is an example of sets of identical proteins forming cage-like spherical shells?
-protein coats of viruses
68
what are examples of mixtures of various proteins and RNA/DNA?
-ribosomes (proteins + rRNA)
-viruses (capsid proteins w/RNA or DNA)
-when dissociated into their constituent macromolecules and then mixed back together, they will reassemble into their original form
-shows the self-organization aspect of a cell
69
what is a globular protein?
-most discussed shape
-ex: enzymes
-ball with an irregular surface containing multiple subunits
70
what are fibrous proteins?
-relatively simple elongated 3D structures (need to span a long distance)
-ex: alpha keratin, intermediate filaments, extracellular matrix proteins
71
what are the characteristics of alpha keratin?
-dimer of two identical subunits wound into a coiled coil
-extremely stable and long lived
-found in hair, nails, and horns=
72
what are the characteristics of intermediate filaments?
-rope-like components of the cytoskeleton
-gives the cells mechanical strength (skin cells)
73
what are the proteins abundant in the extracellular matrix?
-collagen
-elastin
74
what is the purpose of the extracellular matrix?
-binds cells together to form tissues
-proteins within often are secreted by the cells into the surroundings where they assemble into sheets or long fibrils
75
what are the characteristics of collagen?
-most abundant in animal tissues
-3 polypeptide chains in a helix with the non-polar amino acid glycine at every 3rd position at its core (due to vdw and hydrophobic force
-form collagen fibrils (overlapping arrays of collagen through side by side and end to end binding)
-fibrils are very strong and help to hold tissues together
76
what are the characteristics of elastin?
-loose and unstructured polypeptide chains covalently linked into an elastic meshwork
-enables skin, arteries, lungs, etc to stretch and recoil without tearing
-able to do so through the individual protein molecules ability to uncoil reversibly when stretched
77
what is the purpose of covalent cross linkages within proteins?
-help proteins maintain structure when moving in and out of the cell between the differing conditions
-tie together amino acids within the same polypeptide chain or join together many polypeptides in a larger complex (ex: collagen fibrils and elastin fibres)
78
what is an example of a common covalent cross linkage?
-disulfide bridges
79
what are disulfide bridges?
-linking together two SH groups from adjacent cysteine side chains (within the ER, do not form in the cytosol due to reducing agents present)
-only present in secretory proteins
80
do disulfide bridges change a proteins conformation?
-do not change a proteins conformation, but instead act as an "atomic staple" to reinforce the conformation
81
what is a universal function of proteins?
-binding other molecules
-biological properties depend on this physical interaction
82
what are examples of protein binding?
-ex: antibodies bind viruses, bacteria, white blood cells
-ex: enzymes bind substrates (go through chemical rxns, hexokinase binds ATP and glucose)
-ex: actin binds to each other to form a filament
83
what are characteristics of protein binding?
-binding can be tight and long-lived or weak and short-lived depending on the need (needs weak bonds between, number of weak bonds determines strength)
-binding has specificity (binds only 1 or a few molecules)
-binding regulates protein activity
-binding can attach the protein to a particular location in the cell (cell membrane, cytoskeleton, extracellular matrix)
84
what is a weak and short-lived example of protein binding?
-enzyme and their substrate
85
what is a strong and long-lived example of protein binding?
-actin binding together to form filaments
86
what is a ligand?
-substance that is bound by a protein
-can be an ion, organic molecule, macromolecule, etc
87
what is a binding site?
-region of a protein that associates with a ligand
-typically a cavity on the protein surface where the ligand fits that is formed by a particular arrangement of AA side chains
88
what is a substrate?
-the ligand that binds an enzyme and is converted into chemically modified products
89
what is an active site?
-binding site where a substrate binds and catalysis takes place
90
what is a transition state?
-the conformation of the enzyme-substrate complex when it is distorted to lower the activation energy
91
what is a metabolic pathway?
-network of enzymatic reactions where the product of one reaction becomes the reactant for another
-many drugs inhibit enzymes by stopping a specific metabolic pathway (stopping the production of something)
92
what is the first step to enzyme function?
-ligand binding
93
do enzymes need an input on ATP?
-NO
94
what do many proteins need in order to perform their function?
-the help of small nonprotein molecules
-cofactors or coenzymes
95
what is a cofactor?
-small INORGANIC molecule that aids enzymes
-ex: heme groups use iron in hemoglobin
-ex: carboxypeptidase cuts polypeptide chains with the help of zinc
96
what is a coenzyme?
-small ORGANIC molecule that aids enzymes
-ex: biotin aids enzymes in transferring carboxyl groups
-ex: retinal aids rhodopsin in absorbing light in our eyes
97
where do we get many of our coenzymes?
-in our diet
-many coenzymes are vitamins (retinal is vitamin A)
98
what controls the amount of protein within a cell?
-gene expression (make or dont make)
-rate of protein degradation (survive or degrade)
99
how is protein activity controlled?
-confine proteins to sub-cellular compartments (ex: digestive enzymes in lysosomes)
-adjust activity at the level of the protein itself through regulatory sites
100
what are regulatory sites?
-site where a molecule binds to alter the rate at which an enzyme functions (on/off / inhibit/activate)
101
what does the control with regulatory sites depend on?
-protein interactions with other molecules
-change in conformation often results (structure = function)
102
does each enzyme have its own substrate?
-not necessarily
-multiple enzymes can complete for the same substrate
103
what is feedback inhibition? what type of regulation is this?
-an enzyme acting early in a reaction pathway is inhibited by a molecule produced later in that pathway
-when a product accumulates it slows down the catalytic action on the original substrate
-negative regulation (prevents the enzyme from acting)
104
what is positive regulation?
-product in one branch of the metabolic maze stimulates the activity of an enzyme in another pathway
-enzyme is promoted to act
105
what are allosteric proteins?
-proteins that have two different conformations based on what ligands are bound (active and inactive conformations)
-protein will spontaneously switch between these 2 conformations if nothing is bound
-ligand stabilizes the protein in the correct conformation (activating or inhibiting)
106
what is the process of phosphorylation?
-the attachment of a phosphate group to an amino acid side chain (covalently)
-negative charges from the phosphate group interact with positively charged side chains to cause conformational changes in a protein
-the removal of the group returns the protein to its original conformation
-can activate or inhibit a protein depending on the protein itself and the site of phosphorylation
-can create docking sites for other molecules
107
how common is phosphorylation?
-more than 1/3 of the proteins in a mammalian cell are phosphorylated at any one time
108
what is an example of phosphorylation taking place in the cell?
-signal transduction pathways
-uses a protein kinase and protein phosphatase
-protein kinase transfers a phosphate from ATP to a serine/threonine/tyrosine OH group
-protein phosphatase removes a phosphate group (can be specific or broad)
-occurs in a quick loop cycle (turning on and off)
109
what is an example of how receptor tyrosine kinases go through phosphorylation?
-extracellular signals cause these receptor proteins to phosphorylate themselves on certain tyrosines which then serve as docking sites for intracellular signalling proteins to bind and activate which changes the cells behaviour (conformation change occurs)
-found in the cell membrane
110
what are other ways that proteins are modified by additions of other molecules through covalent bonding?
-acetyl groups added to lysine side chains (in histones that form nucleosomes, allow for DNA to loosen)
-fatty acids added to cysteine side chains to associate a protein to the cell membrane (anchoring)
-ubiquitin attaches and brings proteases to the protein to mark it for degradation
111
are proteins only modified at one specific site?
-NO
-many proteins can be modified at many different sites
-ex: p53 protein responds to DNA damage (needs to call upon many other proteins), therefore it can be modified on at least 20 different sites (wide range of behaviour)
112
what can modifications in proteins affect?
-activity
-stability
-binding partners
-location within the cell
113
what are GTP binding proteins?
-GTP bound tightly to a protein
-act as molecular switches
-active conformation is when GTP is bound to the protein (binds fast)
-hydrolyzation of GTP to GDP switches the enzyme to its inactive conformation (GDP dissociates slowly)
-the reactivation is often stimulated by cell signals
-often play a role in intracellular signalling pathways
-reversible process just like phosphorylation
114
what are motor proteins?
-proteins that generate the forces responsible for muscle contractions and other cellular and intracellular movements (moving of organelles and macromolecules)
-present in mitosis and meiosis to move chromosomes to the metaphase plate
-myosin walks along actin filaments
-vesicle transport and movement along the cytoskeleton requires these
-needs ATP so they are also ATPases
-conformational changes are unidirectional and typically consecutive (multiple in a row)
115
how are the conformation changes within motor proteins unidirectional and why?
-want the protein to only move towards the destination
-one step must be irreversible
-ATP molecule binds to the protein
-hydrolysis of the ATP releases a burst of free energy and moves the protein forward
-step is irreversible because it would be energetically unfavourable to add the phosphate back
116
what are protein machines?
-highly coordinated linked sets of proteins
-hydrolysis of ATP or GTP drives an ordered series of conformational changes
-enables the ensemble of proteins to coordinate their movements
-typically successive reactions in a series
-ex: protein synthesis, DNA replication, transcription
117
what are scaffold proteins?
-large molecules that contain binding sites recognized by multiple proteins (bring protein complexes together through rapid collisions )
-enhance the rate of a cell process while confining it to a particular area of the cell
-not permanent structure
-can be rigid or elastic (unstructured regions that bend or sway)
118
what can also act as scaffolds instead of proteins?
-long molecules of RNA
119
what are biomolecular condensates?
-collections of proteins and RNA held together by a continuously shifting set of weak interactions (non-covalent)
-fluid membraneless subcompartments that perform a function
-contain at least one type of RNA or protein scaffold with client molecules bound
-remains separated from its surroundings
-existence can be transient
120
what are clients?
-molecules that bind to the scaffold and become concentrated
121
what do the weak interactions in biomolecular condensates allow for?
-individual molecules to come and go
122
what is an example of a biomolecular condensate?
-nucleolus
-proteins and rRNA are the clients
-site of ribosome synthesis
-transient existence (disappears in phases of mitosis and meiosis)
123
What are the steps to protein purification?
-Take tissues/cells and break them open to release their contents
-use a centrifuge to complete fractionation
-chromatography
-electrophoresis
124
what is the mixture of all the cells/tissues contents referred to as?
-cell homogenate or extract
125
how are the cells/tissues broken open?
-using a detergent or using sonication (sound waves)
126
what is fractionation?
-separating out the class of molecules of interest
-typically using a centrifuge or ultracentrification (can seperate the membrane)
127
what is chromatography?
-separating individual components into fractions based on the properties of the protein (size, shape, charge)
-affinity chromatography
128
what is affinity chromatography?
-separates polypeptides based on their ability to bind particular molecules (properties)
-a protein is attached to the matrix of an affinity column and mixtures of protein are then applied to the column
-some proteins will binds and others will pass through the column
-proteins that bound are then eluted with high salt or a change in pH to have a sample of purified proteins
129
what is electrophoresis?
-polypeptides migrating through a gel at different speeds depending on their size and net charge
-big proteins will move slower
-small proteins will move faster
130
how was protein structure first determined?
-through a direct analysis of amino acids by breaking the protein into smaller pieces using protease
-identities were then determined chemically
-the first protein sequenced this way was insulin
131
what is a faster method of determining protein structure?
-mass spectroscopy
132
what is mass spectroscopy?
-determines the exact mass of every peptide fragment
-allows for identification through a database which applies the knowledge of the genetic code
-blasts peptides with a laser so they become electrically charged and gaseous allowing them to fly towards the detector
-the detector measures the time it takes for them to reach it which is related to mass and charge
133
what are ways of determining the 3D structure of proteins?
-X-ray crystallography
-nuclear magnetic resonance (NMR) spectroscopy
-cryo-electron microscopy (cryoEM)
-all give detailed pictures and data on the shape of a protein
-combine these pictures with knowledge of amino acids sequence and you can figure out the position of each atom in the molecule
-takes a LONG time (AI now helps a lot)
134
what is x-ray crystallography?
-the final protein is taken and crystallized
-x-rays are then blasted which bounce off the protein based on e- density
-watson and crick figured out DNA structure through Rosalind Franklins work with this
135
what is NMR spectroscopy?
-magnets that push and pull on different functional groups to determine which atoms are present
-adds knowledge to the structure
-if the protein is too big it can be overwhelming and complex
-commonly done in conjunction with mass spectroscopy
136
what is cryoEM?
-transmission electron microscope
-protein is fixed in an ethanol ice and pictures are taken on every face to find out the shape (bulges + binding sites)
-a computer makes it into a 3D shape
-expensive
137
why do we care about determining protein sequences and structures?
-because we want to develop a basic understanding of how cells operate
-very important for human health
-knowing the structure can allow us to design new drugs to alter metabolic pathways or stop infections
138
how has AI helped us with determining protein structures?
-AI is used to learn how proteins fold by analyzing readily available data
-can predict the structures of approximately half of all proteins
-can understand sequence patterns
139
what are sequence patterns?
-prove that most proteins belong to families
-the same domain between proteins will play the same role no matter what cell it came from
140
how do we use genetic enginnering?
-make new proteins and enzymes that perform unusual tasks (metabolizing toxic wastes, synthesizing life-saving drugs)
-a synthetic protein that contains a cage that can swing open like a latch when exposed to the key compound was developed
-typically for the delivery of drugs or specific molecules to switch on a target gene, boost immune response, or trigger cell death
141
what therapeutic proteins do bacteria, yeast, and mammalian cells mass-produce?
-insulin
-human growth hormone
-fertility enhancing drugs
BIOL2520
flashcards
Decks in class (19)
# Cards
-
Topic 1 - Introduction to Cell Biology80
-
Topic 2 - Chemical Components of Cells80
-
Topic 3 - Bioenergetics, Enzymes, and Metabolism54
-
Topic 4 - Protein Structure and Function141
-
Topic 5 - DNA and Chromosomes66
-
Topic 6 - DNA Replication and Repair87
-
Topic 7 - From DNA to Protein116
-
Topic 8 - Control of Gene Expression78
-
Topic 9 - Genetic Evolution and the Study of Genes56
-
Topic 10 - Membrane Structure51
-
Topic 11 - Membrane Transport54
-
Topic 12 - Cell signalling39
-
Topic 13 - Mitochondria and Energy Generation36
-
Topic 14 - Cell Division51
-
Topic 15 - Control of Cell Growth, Division, and Survival24
-
Topic 16 - Sexual Reproduction17
-
Topic 17 - Cell Connections0
-
Topic 18 - Stem Cells0
-
Topic 19 - Cancer0
